[Participation of 2-C-methyl-D-erythritol-2,4-cyclopyrophosphate in reactions of bacteria on oxidative stress and their persistence in macrophages]. / Uchstie 2-C-metil-D-éritritol-2,4-tsiklopirofosfata v reaktsii bakterii na okislitel'nyi stress i v ikh persistentsii v makrofagakh.

In aerated medium, Corynebacterium ammoniagenes cells accumulate 2-C-methyl-D-erythritol-2,4-cyclopyrophosphate (MEC) during heat shock and in the presence of O2--generating compounds or ozone. The ability to accumulate MEC was genetically transformed from C. ammoniagenes to E. coli XL-1; transformed E. coli (2-31 clone) accumulates MEC in the presence of glucose and glucose oxidase (generation of H2O2) or benzylviologen (generation of O2-); the viability of transformed bacteria inside the murine peritoneal macrophages also significantly increases. However, model conditions of phagosomes of warm-blooded animals (NO + H2O2 + O2-) did not cause MEC accumulation by C. ammoniagenes but increased the formation of polyphosphate which can be due to selective oxidative aberration of biosynthetic processes. Growth rate of Acanthamoeba castellanii on solid medium with bacterial lawn was not significantly different in C. ammoniagenes, C. ammoniagenes with preaccumulated MEC, E. coli XL-1, and E. coli 2-31 and did not depend on the accumulation of MEC by bacteria. Unlike the recipient E. coli strain, the transformed 2-31 clone synthesizes two nonpolar lipids (Rf = = 0.85 and 0.75; TCL on Silufol in hexane) and carotinoid pigments; this can be due to changes in metabolic pathways of isopentenylpyrophosphate that can be a precursor of MEC biosynthesis. Thus, MEC is involved in bacterial responses to certain components of oxidative stress and in bacterial persistence inside the macrophages.

The ability of a recombinant Escherichia coli strain to synthesize 2-C-methyl-D-erythritol-2,4-cyclopyrophosphate correlates with its tolerance to in vitro induced oxidative stress and to the bactericidal action of murine peritoneal macrophages.

A number of bacteria are able to synthesize 2-C-methyl-D-erythritol-2,4-cyclopyrophosphate (BOSS) in response to oxidative stress. Here we show that the ability to synthesize BOSS can be genetically transferred from Corynebacterium ammoniagenes to Escherichia coli. A total DNA library from C. ammoniagenes ATCC 6872 established in the pBluescript SKII+vector backbone was transfected into E. coli XL-1 blue. Recombinant clone 2-31, which was resistant to redox-cycling agents, was selected. NMR studies showed that this clone was able to synthesize BOSS. We also studied the resistance of clone 2-31 to the bactericidal action of macrophages. Clone 2-31 cells had better survival within murine peritoneal macrophages than parental E. coli XL-1-blue cells. Since the ability to synthesize BOSS correlates with increased survival of bacteria within macrophages, we suggest that the pathogenicity of Corynebacteria could be mediated through the synthesis of BOSS.

[Effect of antiseptics and redox-cycling agents on Corynebacterium ammoniagenes and related microorganisms in relation to synthesis of a new macroergic compound]. / Deistvie antisepticheskikh sredstv i redokstsikliruiushchikh agentov na Corynebacterium ammoniagenes i rodstvennye mikroorganizmy v sviazi s sintezom novogo makroérga.

Sublethal concentration of the antiseptic composition Desoxon-1 was shown to provoke in cells of Corinebacterium ammoniagenes in a liquid medium the biosynthesis and accumulation of a novel macroergic 2-methylbutane-1,2,3,4-tetraol-2,4-cyclopyrophosphate. This substance is also synthesized when C. ammoniagenes is cultivated in a solid agar medium supplemented with benzylviologen. Cells preloaded with the new cyclopyrophosphate maintain its content when treated with 4% phenol, DP-2, Desoxon-1 or boiled and heated in an autoclave. Experiments with Mycobacterium tuberculosis and BCG revealed the ability of these bacteria to grow in a medium supplemented with BV++ possibly due to ability of synthesis of a new cyclopyrophosphate which was shown to correlate with resistance toward redox-cycling drugs. Accumulation of polyphosphates in the control cells of M. tuberculosis was illustrated by 31P-NMR spectroscopy and disappearance of the polyphosphates during cultivation in a BV(++)-supplemented medium. No signal of the new cyclopyrophosphate was yet registered in cells of M. tuberculosis by 31P-NMR.